Abstracts - Society for Developmental Biology

137
Henkels, Julia; Zamir, Evan, Georgia Institute of Technology, Atlanta, United States
During embryonic development, the primitive streak (PS) is the organizing center of gastrulation, an essential process that
results in the formation of the three germ layers and requires complex biochemical and mechanical spatiotemporal
signaling. Current models for PS formation and gastrulation largely ignore the biomechanical environment. Here, for the
first time, we present biomechanical properties of the early avian embryo, a classical model for studying gastrulation. To
test our hypothesis that the spatiotemporaldifferences in stiffness were due to actomyosin contraction, we inhibited
actomyosin contractility via the Rho kinase (ROCK) pathway using the small-molecule inhibitor Y-27632. Explants were
taken from pre- and post-PS embryos and indented using an atomic force microscope. Electroporation and time-lapse
microscopy combined with individual cell tracking provided cell convergence velocities. Treated samples were exposed to
100 mM Y-27632 for 1 h. ROCK-mediated actomyosin contraction appears to be essential for PS formation, the
convergence of cells to the PS during gastrulation, and the increased stiffness of the PS relative to pre-PS stages and other
regions of the post-PS embryo. Further, we found that regions of the embryo outside the PS soften relative to the pre-streak
embryo. These data suggest that genetically-regulated tissue relaxation may be as essential as contraction during
morphogenesis. Future models of the fundamental early morphogenetic movements should account for 1) the critical role
of Rho kinase-mediated actomyosin contractility and 2) the significance of tissue softening as well as contraction to guide
development.
Program/Abstract # 416
PCP pathway controls polarized actomyosin localization through septin 7 during collective cell movements
Shindo, Asako; Wallingford, John, University of Texas at Austin, Austin, United States
The Planar Cell Polarity (PCP) pathway is a critical regulator of cell behaviors during development. Convergent Extension
(CE) is an essential collective cell movement regulated by the PCP pathway. Although PCP signaling is necessary for
establishing cell polarity during CE, little is known about how it controls the cytoskeletal machinery, such as actomyosin,
that execute cell behaviors. We have previously shown that septins have a role in controlling cortical stability downstream
of PCP signaling. In the present study, we show that septin 7 and F-actin accumulate at the medio-lateral cell cortices.
While, activated actomyosin, as assessed by phosphorylation of myosin regulatory light chain (pMYL), localizes at the
antero-posterior cell cortex. Septin 7 KD disrupts these polarized F-actin and pMYL localizations, as we observed ectopic
accumulation at the cortices. Additionally, pMYL protein level is increased in septin 7 KD cells. Finally, we find that
septin 7 is required to maintain polarized cortical forces, which enable cells to intercalate between each other along the
medio-lateral axis. From these results, we conclude that polarized actomyosin distribution is regulated by the PCP pathway
through septin 7 to generate the directional force driving collective cell movement.
Program/Abstract # 417
Uncovering the function of TMED2 during trophoblast differentiation
H, Taghreed; Abeer Zakariyah, Loydie A. Jerome-Majewska, McGill University, Montreal, Canada
Transmembrane emp24 domain trafficking protein 2, (TMED2) is a member of the p24 family of proteins involved in
vesicle transport between the ER and Golgi. During vesicular transport between the ER and Golgi p24 proteins function as
receptors for both cargos and coat proteins. Our group showed that Tmed2 is required for normal embryo and placental
development in mouse and that syncytiotrophoblast cells of the mouse labyrinth placenta failed to differentiate in
homozygous mutant embryos. In human placenta, we showed expression of TMED2 between 5.5 and 40 weeks of
gestation in all trophoblast cell types. We noted that early in gestation TMED2 was more highly expressed in
cytotrophoblast cells versus syncytiotrophoblast. The choriocarcinoma cell lines BeWo and JEG-3 are widely used for the
study of trophoblast differentiation. These cells share many properties with villous trophoblast in terms of their
morphology, biochemical markers, and hormone secretion. We found that TMED2 was more highly expressed in a
choriocarcinoma cell line, BeWo, which can be induced to differentiate and form syncytiotrophoblast when compared to
the JEG-3 cell line, which does not fuse to form syncytiotrophoblast. We hypothesized that TMED2 is required for fusion
of trophoblast cells during syncytiotrophoblast differentiation. To test this hypothesis we are examining the function of
TMED2 during trophoblast differentiation of BeWo and Jeg-3 cell lines. We will show our plans to ectopically express
TMED2 in Jeg-3 cells and to knockdown TMED2 expression in BeWo choriocarcinoma cells with shRNA. Our work
suggests that TMED2 is required for trafficking cargoes that are essential for placental development.
Program/Abstract # 418
Epithelial intercalation drives elongation of the mouse neural plate
Williams, Margot L.K.; Yen, Weiwei; Lu, Xiaowei (University of Virginia, Charlottesville, United States); Lewandoski,